Nilsen Timothy W
Cold Spring Harb Protoc. 2013 Jun 1;2013(6):579-83. doi: 10.1101/pdb.prot075176.
HeLa cells are the archetypal tissue culture cell line for the preparation of mammalian cell-free systems. These cells, derived from a cervical carcinoma, have been maintained in culture since the late 1940s. They grow with a doubling time of ∼24 h and can be cultured in medium containing fetal bovine serum (optimal serum for growth) or medium containing horse serum (in which they grow more slowly, but this serum is considerably less expensive). Nuclear extracts prepared from these cells have been used to determine the mechanisms of splicing and polyadenylation, and such extracts have been characterized extensively. HeLa cells are usually the cell type of choice for initiating cell-free analysis of nearly any aspect of mammalian gene expression. In some instances (e.g., analysis of tissue-specific alternative splicing), it is necessary to use nuclei from a different cell type. We have found that the protocol described here can be used successfully to prepare active nuclear extracts from a wide variety of tissue culture cells, including Drosophila S2 cells.
HeLa细胞是用于制备无细胞哺乳动物系统的典型组织培养细胞系。这些源自宫颈癌的细胞自20世纪40年代末以来一直在培养中保存。它们的倍增时间约为24小时，可以在含有胎牛血清（生长的最佳血清）的培养基中培养，也可以在含有马血清的培养基中培养（在这种培养基中它们生长较慢，但这种血清要便宜得多）。从这些细胞制备的核提取物已用于确定剪接和聚腺苷酸化的机制，并且对这类提取物进行了广泛的表征。HeLa细胞通常是启动对哺乳动物基因表达几乎任何方面进行无细胞分析时的首选细胞类型。在某些情况下（例如，组织特异性可变剪接分析），有必要使用来自不同细胞类型的细胞核。我们发现，此处描述的方案可成功用于从多种组织培养细胞（包括果蝇S2细胞）制备活性核提取物。
